In most eucaryotic genes, a common set of nucleotides has been found before the start site of RNA transcription. These nucleotides, referred to as either a TATA or Goldberg-Hogness box, are important determinants of transcription in vivo and in vitro. The control region for the late SV40 transcripts have been examined by site specific mutagenesis and by generation of deletion mutations. Mutations that enhance or suppress the transcriptional activity of a single start site have been identified and differ from the consensus Goldberg-Hogness box. An additional cluster of nucleotides that is located at base positions 75-96 also control the level of transcription. It maintains this enhancing effect when its position relative to the 5 foot start site is altered.